Breast cancer is the most common cancer in women is often associated with high morbidity and mortality rates, with great financial impact on health system. Besides, the disease is characterized by the high rates of metastasis, which worsen significantly the prognosis. This process is associated with two regulatory molecules, microRNAs (miR), specially miR-17, and ROCK-1, which overexpression has been associated to tumor growth and metastasis. In contrast, melatonin has shown oncostatic and anti-metastatic properties by reducing the cell ability to migrate and invade the tissue, besides the inhibition of cell proliferation. The aim of this study was to investigate the effect of melatonin to modulate miR-17 and ROCK-1, a possible candidate gene to miR-17 target in metastatic breast cancer cell line, MDA-MB-231. To determine the effect of melatonin to modulate miR-17 and ROCK-1, MDA-MB-231 cells were treated with melatonin and anti-miR-17-5p. ROCK-1 and miR-17 gene expression were accessed by real time PCR and ROCK-1 protein expression verified by immunocytochemistry and western blotting. Migration and invasion assay was performed to verify the action of melatonin and anti-miR-17-5p to inhibit these processes. In the in vivo study, was developed pulmonary metastasis model followed for six weeks, the tumor induction was continued for 4 weeks, and treatment with anti-miR-17-5p inhibitor for two more weeks. At the end of treatment, animals were euthanized, the lungs removed and used for analysis of miR-17-5p and Let-7c (positive control) and ROCK-1 gene expression. ROCK-1 protein was analyzed by immunohistochemistry. MiR17-5p inhibition managed directly modulate gene and protein expression of ROCK-1 in MDA-MB-231 cells, as well, the gene expression of MYC. In additional, the migration and invasion were decreased after melatonin and anti-miR-17-5p treatment. To validate the findings of the study in vitro with miR-17, was used for lung metastasis model in athymic nude mice. According to our findings, normal animals without metastasis (negative control) had lower levels of miR-17 compared to animals with metastasis and without treatment (positive control). In contrast, animals with metastasis who received anti-miR17-5p treatment, interestingly also had low miR-17 levels compared to positive control animals. Furthermore, it was observed fewer metastases and a reduction in ROCK-1 protein expression in these treated animals compared to positive control animals. Our results demonstrated that miR-17-5p inhibition can modulate ROCK-1 gene and protein in MDA-MB-231 cells and decreasing the number of lung metastases in treated animals. Furthermore, melatonin can act as a synergic mechanism to decrease migration and invasion on metastasis processes mediated by ROCK-1. Citation Format: Borin TF, Pongeluppi RI, Gelaleti GB, Leonel C, Moschetta MG, Ferreira LC, Zuccari DAPdC. Modulatory action of melatonin and miR-17 on ROCK-1 in breast cancer metastasis model. [abstract]. In: Proceedings of the Thirty-Eighth Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2015 Dec 8-12; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2016;76(4 Suppl):Abstract nr P2-03-02.

Background: In recent years, inhibition of EGFR signaling has emerged as new treatment strategy for oral squamous cell carcinoma (OSCC).The EGFR-directed inhibitor cetuximab is the only approved targeted therapy for the treatment of OSCC.The EGFR status may influence the response to cetuximab treatments.Material and Methods: The subjects were 24 patients with primary OSCC who underwent surgical resection at the Department of Oral and Maxillofacial Surgery at Kanazawa University Hospital between 1998 and 2008.And, three human oral squamous cell carcinoma cell lines established from tumor biopsies with different grade of invasive abilities were used: OSC-20 (low grade invasive cells), OSC-19 (low grade invasive cells) and HOC313 (high grade invasive cells).Results: In this study, by analyzing the immunomarker for EGFR, we found that it indicated positivity in 58.3% of all cases we analyzed, and the invasiveness was inversely correlated with the expression of EGFR.We quantified the expression level of EGFR and analyzed the correlation between EGFR expression and cetuximab sensitivity using three different grade invasive human OSCC lines.The EGFR expression in high grade invasive cells was significantly down-regulated compared to low grade invasive cells.There was no significant anti-proliferative effect in high grade invasive cells treated with various concentrations of cetuximab.The EMTassociated genes N-cadherin, vimentin and Snail were up-regulated in high grade invasive cells.Low grade invasive cells displayed characteristics of typical epithelial cells, including expression of E-cadherin and absence of N-cadherin, vimentin and Snail.TGF-beta induced this low grade invasive cell to undergo an EMT-associated gene switch that results in low EGFR expression.Conclusions: These data suggested that loss of EGFR expression in OSCC was associated with EMT, and might have functional implications in tumor invasion and resistance to cetuximab treatments.No conflict of interest.

Meeting abstracts Interleukin (IL)-25, a cytokine active in inflammatory processes and production of proinflammatory cytokines, is secreted by mammary epithelial non-malignant cells and induces apoptosis in tumour cells by differential expression of its receptor IL-17RB, highly expressed in

Caveolin-1 (Cav-1) is a structural protein present in invaginations of the cell membrane. In human breast cancer, the cav-1 gene is believed to be a tumor suppressor gene associated with inhibition of tumor metastasis. However, little is known about its expression, regulation and function in canine mammary tumors. Expression levels of cav-1 were investigated using real- time PCR and immunohistochemical detection with an anti-human Cav- 1 antibody. Gene expression stability of different samples was analyzed using the geNorm software. Mammary tumors from 51 female dogs were compared to normal mammary tissue from 10 female dogs. Malignant mammary cells showed a loss of Cav-1 expression by quantitative RTPCR and weak Cav-1 staining by immunohistochemistry compared to normal mammary gland tissue. There was a significant relationship between outcome and immunostaining as well as with tumor size, indicating that caveolin subexpression has a positive predictive value and is related to higher survival and smaller tumor size. Our findings indicate that Cav-1 is a potential prognostic marker for canine mammary tumors.

S215questionable and still debated.Within relevant time intervals in both natural and clinical course of disease, we analyzed quantitative ER values in order to determine cut points, at minimal p value, allowing us to distinguish patients subgroups at different risk for relapse.Material and Methods: Study included 228 postmenopausal breast cancer patients where 102 pN0 patients received no adjuvant therapy and 126 patients received adjuvant tamoxifen treatment.ER concentrations were measured in breast cancer tissue extracts by use of classical biochemical method.Within the relevant time intervals (0-2.5, 2.5-5 and 5-12 years), all patients had equal, maximal follow-up.Survival curves for disease-free interval (DFI) were estimated according to method of Kaplan and Meier and compared by log-rank test.Results and Discussion: In most studies, analyses are based on diverse follow-up intervals, having various median follow-up times.In our study, all relaps-free patients were monitored during whole folow-up periods, relevant for occurrence of distant metastases.For defining ER status in breast cancer, not only that different arbitrary values are used as ER cut-off values but analysis of ER quantitative values is neglected due to use of immunohistochemistry. Analysis revealed ER concentration of 52 fmol/mg (p = 0.02) as cut point for pN0 patients indicating prognostic relevance of ER only within follow-up period of 5-12 years after diagnosis.Among patients treated with adjuvant tamoxifen, ER showed predictive value within periods of 0-2.5 (cut-off value at 4 fmol/mg, p = 0.002) and 5-12 years after diagnosis (cut-off value at 158 fmol/mg, p = 0.05) that might be important in planing of enocrine therapy.Conclusions: Within different, relevant intervals of natural as well as clinical follow-up, there is no unique cut-off value for ER expression in breast cancer.Within same, relevant intervals of natural and clinical follow-up, cut-off values for ER expression in breast cancer are not equal.

Faculdade de Medicina de Sao Jose do Rio Preto (FAMERP), Dept Mol Biol, Sao Jose do Rio Preto, SP, Brazil